Extended Data Fig. 6: MAVS but not c-GAS-STING mediates IFN activation upon Ku degradation.
From: Ku limits RNA-induced innate immunity to allow Alu expansion in primates
a, Western blotting shows that cGAS protein is not expressed in HCT116 cells with Hela cells as the positive control. b, Ku is required in human cells regardless of cGAS expression levels. The mRNA levels of cGAS, a dsDNA sensor provided by the Dependency Map database, were plotted against the XRCC5/Ku80 Gene deletion effect. Effect score =0 (neutral) and <−1 (essential). c-g, Quantitative RT-PCR analyses of IFN targets, ISG15 (c), IFIT1 (d), IFNβ (e), STAT1 (f), and NF-kB target GADD45B (g) validated the critical role of MAVS in mediating IFN activation following Ku-degradation. Parental HCT116 of the AID system cells (n = 9), three independently derived Ku80-AID clones (n = 18 in (c) and (f), 27 in (d-e) and (g)) and three independent MAVS KO clones (n = 18 in (c) and (f), 27 in (d-e) and (g)) were analyzed individually and plotted together. Poly I:C transfection (100 ng/ml, 24 hr) was used as a positive control (n = 9). Bars indicate the mean ± SEM from at least three biologically independent samples. Two-way ANOVA test with Sidak’s correction for multiple comparisons: p < 0.0001 ****; p > 0.05 n.s. h, Representative western blotting analyses of IFN pathway markers in multiple independently derived MAVS, RIG-I, and MDA5 KO clones. Additional clones can be found in Fig. 3a. i, Western blotting analyses of dsRNA sensors RIG-I, MDA5, MAVS, and IRF7 in HCT116, HEK293 and control Hela cells.
